Cancer Drug Bio-factories: Hairy Roots

The Ri (root-inducing) plasmid of Agrobacterium rhizogenes carries agropine genes. When A. rhizogenes infects a plant, a portion of the Ri plasmid DNA enters the host plant cell and causes the production of hairy roots at the site of action. A foreign gene could be inserted into modified Ri plasmid and the recombinant DNA (plasmid) [...]

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Maize Mini-chromosomes. Algae as Biofuel

This post is about two recent developments in the field of biotechnology and environmental science.
1) Agrobacterium-mediated transformation is the method of choice for inserting foreign genes into the plant genome. Through this method one gene could be introduced at a time in a single experiment. However, to introduce two foreign genes, one needs to develop [...]

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Integrating T-DNA into plant genome

After its entry into the plant cell the ssT-DNA complex crosses the nuclear membrane to enter the nucleus and subsequently integrates into the plant genome. VirD2 and VirE2 are the two most important proteins required for this process. VirF also plays a minor role in the same (Hooykaas, P.J.J. and Shilperoort, R.A. (1992). Agrobacterium and [...]

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T-DNA transfer contd…

The T-DNA transfer process starts with the formation of a nick at the right 25 bp repeat. This provides the priming end for synthesis of a DNA single strand. Synthesis of new strand displaces the old strand. The old strand is used in the transfer process: The old strand is transferred to the nucleus. The [...]

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T-DNA transfer contd…

ssT-DNA-protein complex is transferred to the plant nucleus, and during the translocation it passes through three membranes, the plant cell wall and cellular spaces. The ssT-DNA-VirD2 complex formed after the cleavage is coated by the 69 kDa VirE2 protein. The VirE2 protein is a single strand DNA binding protein. It prevents the DNA from the [...]

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T-DNA transfer contd…

The T-DNA is flanked by 25 bp direct repeats. These repeats differ at only two positions between the left and right ends. Mutation or deletion studies (Hille, J., Wullems, G. and Schilperoort, R.A. (1983). Non-oncogenic T-region mutants of Agrobacterium tumefaciens do transfer T-DNA into plant cells. Plant Molecular Biology 2: 155-163) of [...]

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T-DNA transfer

virD is an operon of 4 genes. Out of these four genes two code for VirD1 and VirD2 proteins. VirD1 and VirD2 play a key role in generating T-DNA transfer complex: They recognize the T-DNA border sequences and nick the bottom strand at each border by their endonuclease activity. VirD2 remains covalently attached to [...]

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vir activation

The virA and virG genes are constititively expresssed and respond to a change in the plant by inducing the other genes. VirA is a transmembrane dimeric protein which detects small phenolic compounds, like acetosyringone or alpha-hydroxyacetosyringone, released from wounded plants. The signals for VirA induction, in addition to small phenolics, also include some monosaccharides which [...]

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vir Regulon

The vir (virulence) region lies outside the T-DNA on the Ti plasmid. The 30-40 kb vir region is a regulon of 8 operons. Out of these 8 operons, 6 are essential and 2 are non-essential. The essential operons are virA, virB, virC, virD, virE and virG; the 2 non-essential operons are virF and virH. Each [...]

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What is required during plant transformation?

The transformation of a plant cell requires three types of functions in Agrobacterium:
1) chvA, chvB, pscA: These three loci present on Agrobacterium chromosome are needed at the initial stage of binding the bacterium to the host plant cell. “They are responsible for synthesizing a polysaccharide on the bacterial cell surface”.
2) vir region: The vir (virulence) [...]

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